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To test the therapeutic efficacy of azithromycin (AZM), a macrolide antibiotic for prolonging murine “high-risk” corneal allograft survival.Fully major histocompatibility complex–mismatched corneas were transplanted from C57BL/6 donors to BALB/c recipients with suture-induced vascularized high-risk corneal beds. Recipient mice were either not treated or treated with topical AZM, oral AZM, or both. Evaluation of graft vascularization and clarity was performed in a masked fashion. Lymph nodes were excised and analyzed for CD4, FoxP3, and CD44 by flow cytometry, and for T-cell priming by proliferation and cytokine production in mixed lymphocyte cultures. Corneal whole mounts were evaluated by confocal microscopy.The incidence of graft rejection in the control group (81.8%) was significantly reduced by AZM treatment (18.2% topical, 21.7% oral, 33.3% topical + oral), although corneal vascularization was not affected by the treatment. The frequency of corneas that retained complete clarity after transplantation was higher in the AZM-treated groups. Reduced graft rejection in the AZM-treated groups was not associated with a reduced allospecific T-cell response or increased frequency of regulatory T cells.AZM is effective in prolonging survival of high-risk corneal allografts by an as yet undefined mechanism that does not seem to involve modulation of corneal neovascularization or allospecific T-cell priming.